Human Complement Receptor type 2 (CR2/CR21) is a B lymphocyte receptor for at least three distinct ligands: the iC3b and C3d fragments of complement C3, gp350/220 of the Epstein-Barr virus and CD23 (FceRII). By interacting with these ligands, CR2 plays an important role in modulating the immune response. The ligand binding extracellular structure of CR2 consists of a linear array of 60-70 amino acid-containing modules designated short consensus repeats (SCRs). Proteins containing SCRs comprise a genetically linked family called the regulators of complement activation (RCA). Each RCA protein is involved in binding complement C3 and/or C4 and serving as a receptor or as a complement regulatory protein. Previous studies have shown that the binding site for each of these CR2 ligands is contained either completely (iC3b, C3d and gp350/220) or partially (CD23) within the amino terminal two of sixteen SCRs (SCR 1-2 domain ligand binding sites are related but not identical. Presented herein is a preliminary model of the iC3b binding site that we have recently completed by overlaying apparent iC3b ligand contact residues from CR2 onto the NMR derived coordinates of a double SCR domain from the structurally-related RCA family member, factor H. This analysis shows that the two ligand binding sites on the two SCRs are highly solvent exposed, and that a relative twist is required between the two domains in order to create a hypothetical single surface "patch" on CR2 with which iC3b would bind. Because CR2 interacts within a relatively small region with at least three distinct biologically relevant proteins, we believe that CR2 can serve as an excellent model for SCR-ligand interactions and that a comprehensive and multi-disciplinary approach should be pursued to characterize these binding sites. To that end, we propose three specific aims: 1) Determine by two- dimensional 1H NMR analysis and crystallography the three-dimensional structure of the CR2 SCR 1-2 domain, 2) further characterize and compare the binding sites for each ligand (iC3b, C3d, gp350/220 and CD23) using CR2 derived peptides and mutated forms of recombinant CR2, and 3) identify structurally informative reagents (mAbs and single chain Fv (scFv) antibody fragments from phage display libraries) that selectively inhibit binding of each ligand.